U.S. patent number 6,365,175 [Application Number 09/468,636] was granted by the patent office on 2002-04-02 for petroselinic acid and its use in food.
This patent grant is currently assigned to Unilever Patent Holdings. Invention is credited to Simon Alaluf, Frederick William Cain, Martin Richard Green, Heng Long Hu, Jonathan Richard Powell, Anthony Vincent Rawlings, Julia Sarah Rogers, Allan Watkinson.
United States Patent |
6,365,175 |
Alaluf , et al. |
April 2, 2002 |
Petroselinic acid and its use in food
Abstract
Edible compositions containing petroselinic acid are used for
the preparation of food compositions or food supplements that are
used as anti-inflammatory compositions that inhibit the production
of metabolites of arachidonic acid and/or reduces the formation of
intracellular adhesion molecules or as anti-aging compositions with
a positive impact on wrinkling, sagging, photodamaged skin, dry
skin, flaky skin and age spots.
Inventors: |
Alaluf; Simon (Shambrook,
GB), Green; Martin Richard (Shambrook, GB),
Powell; Jonathan Richard (Shambrook, GB), Rogers;
Julia Sarah (Shambrook, GB), Watkinson; Allan
(Shambrook, GB), Cain; Frederick William (Wormerveer,
NL), Hu; Heng Long (Abbeymead, GB),
Rawlings; Anthony Vincent (Bebington, GB) |
Assignee: |
Unilever Patent Holdings
(Vlaardingen, NL)
|
Family
ID: |
27239595 |
Appl.
No.: |
09/468,636 |
Filed: |
December 22, 1999 |
Foreign Application Priority Data
|
|
|
|
|
Dec 22, 1998 [EP] |
|
|
98310626 |
Dec 22, 1998 [EP] |
|
|
98310627 |
Mar 17, 1999 [EP] |
|
|
99302067 |
|
Current U.S.
Class: |
424/439;
424/78.05; 514/786; 514/787 |
Current CPC
Class: |
A23D
7/001 (20130101); A23D 7/015 (20130101); A23D
9/00 (20130101); A23D 9/007 (20130101); A23D
9/013 (20130101); A23G 3/40 (20130101); A23G
9/327 (20130101); A61K 8/361 (20130101); A61Q
19/00 (20130101); A61Q 19/004 (20130101); A61Q
19/007 (20130101); A61Q 19/02 (20130101); A61Q
19/08 (20130101); A23L 33/12 (20160801) |
Current International
Class: |
A23D
9/007 (20060101); A23D 9/00 (20060101); A23D
7/015 (20060101); A23D 7/00 (20060101); A23D
9/013 (20060101); A23G 3/00 (20060101); A23G
9/32 (20060101); A23L 1/30 (20060101); A61K
047/00 (); A61K 031/74 () |
Field of
Search: |
;424/439,78.05
;514/473,557,786,787 |
References Cited
[Referenced By]
U.S. Patent Documents
|
|
|
4918063 |
April 1990 |
Lictenberger |
5242945 |
September 1993 |
Caufield et al. |
5605929 |
February 1997 |
Liao et al. |
6022896 |
February 2000 |
Weinkauf et al. |
|
Foreign Patent Documents
|
|
|
|
|
|
|
777971 |
|
Jun 1997 |
|
EP |
|
0 777 971 |
|
Jun 1997 |
|
EP |
|
777971 |
|
Jun 1997 |
|
EP |
|
61257944 |
|
Nov 1986 |
|
JP |
|
WO 95/00136 |
|
Jan 1995 |
|
WO |
|
Other References
Afifi et al., "Some pharmacological activities of essential oils of
certain umbelliferous fruits", VET. MED. J. GIZA, vol. 42, No. 3,
1994, pp 85-92.* .
Afifi et al., "Some pharmacological activities of essential oils of
certain umbelliferous fruits", VET. MED. J. GIZA, vol. 42, No. 3,
1994, pp 85-92.* .
Yagaloff, "Essential fatty acids are antagonists of the leukotriene
B4 receptor", Prostaglandins, Leukotriens and Essential Fatty
Acids, vol. 52, 1995, pp 293-297.* .
Patent Abstracts of Japan, Publication No. 61181352. .
Yagaloff et al, Prostaglandins Leukotrienes and Essential Fatty
Acids, 52(5):293-297 (1995) (XP 002050601). .
Afifi et al, Vet. Med. J. Giza., 42(3):85-92 (1994) (XP 002050599).
.
Derwent Abstract XP-002120504. .
Derwent Abstract XP-002120503..
|
Primary Examiner: Page; Thurman K.
Assistant Examiner: Fubara; Blessing
Attorney, Agent or Firm: Morgan, Lewis & Bockius LLP
Claims
What is claimed is:
1. An edible encapsulated composition with anti-inflammatory and/or
anti-aging properties comprising a petroselinic acid containing
composition and an anti-oxidant wherein the encapsulating material
is selected from the group consisting of: polysaccharides, sugars,
fats, proteins and amino acids.
2. An edible encapsulated composition with anti-inflammatory and/or
anti-aging properties comprising a petroselinic acid containing
composition and an anti-oxidant according to claim 1 wherein the
encapsulating material is present in an amount of 5-95 wt % on
basis of total encapsulated composition.
3. A method of obtaining at least one effect from the group
consisting of an anti-inflammatory effect that inhibits the
production of metabolites of arachidonic acid and/or reduces the
formation of intracellular adhesion molecules and an anti-aging
effect on skin conditions which comprises orally administrating to
a subject in need of such effect, an edible composition comprising
an effective amount of petroselinic acid together with an
anti-oxidant.
4. The method of claim 3 wherein the composition is a functional
food composition or a food supplement.
5. The method of claim 3 wherein the composition is administered to
obtain a desirable effect on skin conditions selected from the
group consisting of wrinkling, sagging, photodamaged skin, dry
skin, flaky skin and age spots.
6. The method of claim 3 wherein the composition comprises 5-99.9
wt % of fat or fat blend.
7. The method of claim 6 wherein said composition includes a fat
selected from the group consisting of: cocoa butter, palm oil or
fractions thereof, palm kernel oil or fractions thereof,
interesterified mixtures of the above fats, hardened fats or
fractions thereof, sunflower oil, high oleic sunflower oil, soybean
oil, rape seed oil, cotton seed oil, safflower oil, high oleic
safflower oil, maize oil, MCT oils, hardened liquid oils or
fractions thereof and mixtures thereof.
8. The method of claim 3 wherein said composition contains 2-80 wt
% of petroselinic acid.
9. The method of claim 8 wherein said composition contains 5-40 wt
% of petroselinic acid.
10. The method of claim 3 wherein the petroselinic acid is in the
form of its free fatty acid, or a mono-, di- or triglyceride with
at least one petroselinic acid group, a wax ester of petroselinic
acid or a short alkyl ester of petroselinic acid or mixture
thereof.
11. The method of claim 3 wherein said anti-oxidant is selected
from the group consisting of natural or synthetic tocopherols,
natural polyphenols, BHT, BHA, free radical scavengers and enzymes
with anti-oxidant properties.
12. The method of claim 11 wherein said anti-oxidant is a tea
extract.
13. A composition comprising:
(1) a fat composition comprising asymmetric isomers from cis
monounsaturated fatty acid(s), wherein the fat composition
comprises 15-75 wt % of asymmetric isomers from petroselinic acid,
25-50 wt % of saturated fatty acids with 12-24 carbon atoms and
0-60 wt % of isomers of other fatty acids with at least 18 carbon
atoms; and
(2) from 0.01-5 wt % of one or more anti-oxidants selected from the
group consisting of: natural or synthetic tocopherols, natural
polyphenols, BHT, BHA, free radical scavengers and enzymes with
anti-oxidant properties.
14. A composition according to claim 13 wherein the fat composition
(1) comprises three components A, B and C, wherein:
A has a content of asymmetric isomers from monounsaturated fatty
acids of at least 20 wt %;
B has a solid fat content, measured on an unstabilized fat by NMR,
at 20.degree. C. of at least 20, and
C has a content of fatty acids with at least 18 C atoms and a cis 9
double bond of at least 40 wt %,
while components A, B and c are present in amounts of:
15-90 wt % of A,
10-85 wt % of B, and
0-75 wt % of C.
15. A composition according to claim 14 wherein the fat composition
(1) contains an additional component D in an amount of 5-60 wt %,
which component D is an interesterified mixture of fats A and B in
a weight ration of 95:5-5:95.
16. A composition according to claim 13 wherein the fat composition
has a solid fat content unstabilized at 5.degree. C. of 25-85 and
at 35.degree. C. of less than 10.
17. A composition according to claim 14 wherein A is selected from
the group consisting of coriander oil, parsley oil or fungal
oils.
18. A composition according to claim 14 wherein A is a fat obtained
after performing an enrichment in cis 6 isomers of fatty acids of
monounsaturated fatty acids by performing an enzymic conversion on
a fat containing at least 5 wt % of cis 6 isomers of fatty acids of
monounsaturated fatty acids and containing also cis 9 double bonds
using an enzyme specific for cis 9 double bonds and removal of the
products enriched in cis 9 double bonds.
19. A composition according to claim 13 wherein the fat composition
(1) has a trans content of 10-70 wt %.
20. A composition according to claim 14 wherein fat B is selected
from the group consisting of: palm oil stearin, palm oil mid; cocoa
bufter, cottonseed stearin, fully or partially hardened vegetable
oils.
21. A composition according to claim 14 wherein fat C is selected
from the group consisting of sunflower oil, high oleic sunflower
oil, olive oil, bean oil, safflower coil, rape seed oil, palm oil
olein, olein fractions of vegetable oils, high oleic vegetable
oils, corn oil or cottonseed oil.
22. A food product having added thereto a composition according to
claim 13.
Description
FIELD OF THE INVENTION
The invention relates to edible compositions containing
petroselinic acid for anti-inflammatory and/or anti-aging use.
Petroselinic acid is a well known compound that eg is present in
coriander oil in relatively high amounts. Its use in food products
is also disclosed in literature. EP 777971 eg discloses that fat
compositions can be obtained from fats that have high contents of
asymmetric monounsaturated fatty acids, such as petroselinic acid,
ie PSA (>79 wt %) and that have simultaneously low contents of
trans acids (<5 wt %). The fat compositions obtained can be used
as fat replacer with a number of benefits such as providing good
textural properties to the food, while not raising the
LDL-cholesterol levels in the blood serum. When considering example
5, in particular table 3 of this patent application one must
conclude that good product performance is only obtained if the fat
applied contains about 12 wt % PSA. Products based on fats without
PSA perform badly (=control fat 1). Fats with only small amounts
PSA are still grainy, while using a fat with a high content of PSA
(=fat 4 with about 64 wt % PSA via the coriander oil) has a poor
overall appearance. Food products that are indicated are
margarines, dressings, confections, spreads, frozen desserts, ice
cream, mayonnaise, mustards, cheese, dip sauces, bread, biscuit,
dairy products, frying oils, CBE's, candy, meat, egg products, nut
products, vegetable or fruit products, toppings, creams, puddings,
cookies, pastries, pies, crackers, cakes, bread rolls and
ingredients or premixes herefore. As PSA is the active ingredient
that provides the health benefits to the food product there is a
big need to be able to use fats with high levels of PSA but that
perform well when applied in food products. Ie the appearance of
the food products should be good as well.
Inflammation is a key problem in a number of disease states.
Extensive scientific research has focused on identifying compounds
that have anti-inflammatory activity. So far the main treaments are
synthetic compounds in the pharmaceutical area or complex mixtures
of compounds, synthetic or natural. It is known that fish oils can
be used to some extent as an anti-inflammatory. However, fish oils
have the disadvantage of being very unstable and of forming off
tastes very easily. Therefore the use of products based on fish
oils as anti-inflammatory compositions has not found wide
commercial application within the public domain.
U.S. Pat. No. 4,918,063 (1990) discloses that some specific
inflammatory diseases such as peptic ulcer disease or inflammatory
bowel disease can be prevented or treated with mixtures of
saturated or unsaturated phospholipids and saturated or unsaturated
triglycerides and/or sterols. The activity of these mixtures is
attributed to the fact that these mixtures have the ability to
increase or maintain its hydrophobic character by treating the
luminal surface of the gastrointestinal tract. It is expressly
disclosed that the use of unsaturated phospholipids per se does not
work. These compounds only can work in the presence of a
triglyceride or sterol. This thus leads away from using unsaturated
derivatives of fatty acids for the purposes mentioned in this
document. No examples are given for the use of petroselinic acid or
glycerides containing this. The only indication herefore can be
found in a listing in table 1.
The teaching laid down in the above U.S. 063 is confirmed by U.S.
Pat. No. 5,178,873, wherein a method for inhibiting phospholipase
A2 activity is disclosed by using stearidionic acid or a C20:4
unsaturated fatty acid. In the Tables 2 and 3 it is demonstrated in
a comparative example that petroselinic acid is not useful for this
purpose.
Another well recognized consumer problem is formed by the aging of
the skin. In recent years the demand for methods to improve the
appearance of skin and, in particular, for reducing or preventing
the visible signs of wrinkled, aged and/or photodamaged skin has
grown enormously. It is known in the art that the levels of
collagen, a dermal structural protein and decorin, a structuring
proteoglycan in skin are significantly reduced with aged and/or
photodamaged skin (Lavker et al J.Inv.Derm.1979 73:79-66, Bernstein
et al Lab. Invest. 1995 72:662-669). The reduction in these
proteins is associated with a decrease in the tensile strength of
the skin causing wrinkles and laxity.
Extensive research has focused on identifying actives that have
anti-aging activity. One of the main current actives is retinoic
acid, giving wrinkle effacement and dermal repair through boosting
for example by collagen synthesis (e.g. Griffiths et al. N. Eng. J.
med. 1993 329:530-535). According to GB 2 181 349 another solution
was found in the use of triglycerides derived from long chain
polyunsaturated fatty acids in cosmetic or dermatological
compositions. These compositions are thus not edible in general
because they also will contain non edible components. Moreover the
triglycerides derived from these polyunsaturated fatty acids are
unstable in particular with respect to oxygen stability.
We studied whether we could find a natural component that could be
used as anti-inflammatory agent without creating problems of side
effects and not having the negative aspects from fish oils, and
other known anti-inflammatory compositions.
We also studied whether we could find a natural component that
could be used as anti-aging agent without creating problems of side
effects and not having the negative aspects from the known
anti-aging compositions.
We further studied whether we could find fat compositions that
combine high PSA levels with a very good product performance.
The study on anti-inflammatory and antiaging activity resulted in
our finding that petroselinic acid derivatives (ie free fatty acid,
short alkyl ester, short being C1-C4, wax-esters, mono-di- or
triglycerides) can be applied as firstly anti-inflammatory agents
that act on the formation of metabolites from arachidonic acid (a
recognised precursor of inflammatory mediators), or that can reduce
the formation of intracellular adhesion molecules, and thus
contribute to the inflammatory response. Or secondly can be applied
as an anti-aging agent that act by boosting the levels of two
structural proteins in the dermis of the skin, collagen and
decorin. Moreover we found novel fat compositions that combine high
PSA levels with good product performance.
BRIEF SUMMARY OF THE INVENTION
Therefore our invention concerns in the first instance the use of
edible compositions containing petroselinic acid for the
preparation of functional food compositions or food supplements,
wherein the composition containing petroselinic acid is used as an
anti-inflammatory component that inhibits the production of
metabolites of arachidonic acid and/or reduces the formation of
intracellular adhesion molecules.
Our invention further concerns the use of edible compositions
containing petroselinic acid for the preparation of functional food
compositions or food supplements, wherein the composition
containing petroselinic acid is used as an anti-aging component
that boosts decorin levels with a positive impact on skin
conditions selected from the group consisting of wrinkling,
sagging, photodamaged skin, dry skin, flaky skin and age spots.
Functional food compositions being defined as food compositions
containing at least one component with a health benefit. Food
supplements being defined as compositions that are not used as food
per se, but that are used as supplement to the daily food intake,
in general in the form of encapsulated essential food ingredients.
These edible compositions containing petroselinic acid can be
applied in many different forms, but we prefer to apply these
compositions as a composition comprising 5-99.9 wt % of fat or fat
blend.
The fats that can be used in the compositions containing the
petroselinic acid derivative can be selected from the group
consisting of: cocoa butter equivalents, palm oil or fractions
thereof, palm kernel oil or fractions thereof, interesterified
mixtures of above fats, hardened fats or fractions thereof, liquid
oils, selected from sunflower oil, high oleic sunflower oil,
soybean oil, rape seed oil, cotton seed oil, safflower oil, high
oleic safflower oil, maize oil, MCT oils, hardened liquid oils or
fractions thereof and mixtures hereof.
These fats are nearly all natural fats (the exception being MCT
oils which are synthetic fats based on medium chain fatty acids ie
fatty acids with 6-12 carbon atoms). The level of petroselinic acid
in the compositions that we can apply for our purposes can vary
widely, however we prefer to apply petroselinic acid containing
compositions that contain 2-80 wt %, preferably 5-40 wt % of
petroselinic acid derivative (calculated as petroselinic acid).
As already indicated above we found that the petroselinic acid
could be used in different forms. Very good results were obtained
by using the petroselinic acid as free fatty acid, or in a form
wherein it is bound to a glycerol backbone as in mono-di- or
triglycerides. In this latter case the glycerides must contain at
least one petroselinic acid moiety. Other forms are short alkyl
esters of the petroselinic acid, short meaning having one to eight,
preferably 1 to 4 carbon atoms. Also wax esters, i.e. long chain
alcohol esters of petroselinic acid can be applied. Of course also
mixtures of above forms can be used.
We also found that the anti-inflammatory and antiaging effect of
the petroselinic acid can be increased, even in a synergistic way
by using the petroselinic acid derivative in combination with one
or more anti-oxidants. Typical examples of useful anti-oxidants can
be selected from the group consisting of natural or synthetic
tocopherols, natural polyphenols, in particular as present in tea
extracts, BHT, BHA, free radical scavengers and enzymes with
anti-oxidant properties. From these anti-oxidants we prefer the
tocopherols and the polyphenols as present in tea extracts the
most.
The petroselinic acid derivatives will structure the fats when
applied in a fat surrounding. Therefore structured fats comprising
the petroselinic acid composition and wherein the fat has a level
of saturated plus trans fatty acids of 25-75 wt % are also part of
our invention.
Part of the invention are also the blends of the petroselinic acid
containing composition and 0.01-5 wt % of one or more anti-oxidants
selected from the group consisting of: natural or synthetic
tocopherols, natural polyphenols, in particular as present in tea
extracts, BHT, BHA, free radical scavengers and enzymes with
anti-oxidant properties.
Food products or food supplements containing these blends are also
part of our invention. As food products all food products indicated
in EP 777971 could be used.
A very useful form wherein the active petroselinic acid or
derivative can be applied is an encapsulated composition, wherein
the petroselinic acid is encapsulated in an edible encapsulating
material. The encapsulating material is selected from the group
consisting of: polysaccharides, sugars, fats, proteins and amino
acids.
The encapsulating material is present in an amount of 5-95 wt % on
basis of total encapsulated composition.
These encapsulated forms are in general free flowing which makes
the dosing of the food products or food supplements easier.
As a result of our study to find fat compositions that combine high
PSA levels with very good product performance we found that fat
compositions comprising asymmetric isomers from cis monounsaturated
fatty acids, wherein the fat composition comprises 15-75 wt %,
preferably 20-60 wt %, most preferably 30-60 wt % of asymmetric
isomers from cis monounsaturated fatty acids preferably being
petroselinic acid, 25-50 wt %, preferably 30-40 wt % of saturated
fatty acids with 12-24 carbon atoms and 0-60 wt %, preferably 5-50
wt %, most preferably 15-45 wt % of isomers of other fatty acids
with at least 18 carbon atoms, preferably having a cis 9 double
bond, preferably being oleic acid, or linoleic acid, are very
suitable for this purpose.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 illustrates the inhibition of PMA-induced PGE2 production by
petroselinic acid in a fibroblast cell model.
FIG. 2 illustrates the inhibition of PMA-induced ICAM production by
petroselinic acid in a fibroblast cell model.
FIG. 3 illustrates the inhibition of PMA and BSO induced ICAM
production by petroselinic acid and antioxidants in a fibroblast
cell model.
FIG. 4 illustrates the inhibition of PMA-induced ICAM production by
petroselinic acid and genistein in a fibroblast cell model.
FIG. 5 illustrates the inhibition of keratinocyte PGE2 by
petroselinic acid.
FIG. 6 illustrates the protection of keratinocytes in presence of
SDS.
DETAILED DESCRIPTION OF THE INVENTION
According to a preferred embodiment of this aspect of our invention
these compositions comprise three components A, B and C,
wherein:
A has a content of asymmetric isomers from monounsaturated fatty
acids of at least 20 wt %, preferably at least 40 wt %, most
preferably at least 50 wt %
B has a solid fat content, measured on a unstabelized fat by NMR,
at 20.degree. C. of at least 20, preferably at least 45, most
preferably at least 60 and
C has a content of fatty acids with at least 18 C atoms and a cis 9
double bond of at least 40 wt %, while components A, B and C are
present in amounts of:
15-90 wt % of A, preferably 20-75 wt %, most preferably 30-50 wt
%
10-85 wt % of B, preferably 20-75 wt %, most preferably 30-60 wt %
and
0-75 wt %, preferably 5-60 wt %, most preferably 15-50 wt % of
C.
Even more preferred compositions are obtained when the composition
contains an additional fat component D in an amount of 5-60 wt %,
which component D is an interesterified mixture of fats A and B in
a weight ratio of 95:5-5:95. This then results in fat compositions,
wherein the components A,B,C and D are present in amounts of:
5-75 wt % of A
15-60 wt % of B
5-75 wt % of fat C and
5-75 wt % of fat D.
In order to achieve the best mouth feel our fat compositions have a
solid fat content unstabilised at 5.degree. C. of 25-85, preferably
30-50 and at 35.degree. C. of less than 10, preferably less than 5.
These solid fats contents are measured by NMR pulse techniques on a
fat at measurement temperature. The value for the solid fat content
depends on the pretreatment of the fat. Here the solid fat content
is measured on a fat that is not stabilised. I.e. the fat was
melted and kept at 60.degree. C. for 30 min, after which the fat is
cooled to 0.degree. C. and kept hereon for another 30 min,
whereupon the fat is heated to measurement temperature and the
solid fat index is measured after keeping the fat at this
temperature for at least 1 hour.
As a source for the PSA, coriander seed oil is very useful.
Coriander oil can contain up to about 80 wt % of PSA. Other fats
relatively rich in PSA that can be applied as well are: parsley
oil; and a number of fungal oils. PSA can also be obtained from
carrots, fennel: chervil or caraway. The content of cis 6 isomers
of asymmetric monounsaturated fatty acids in fats having
intermediate levels thereof can be increased by performing an
enzymic conversion on a fat containing at least 5 wt % of cis 6
isomers of fatty acids of monounsaturated fatty acids and
containing also cis 9 double bonds using an enzyme specific for cis
9 double bonds and removing the products enriched in cis 9 double
bonds. Enzymes suitable for this conversion are well known, an
example being Geotrichum candidum.
Although the trans content of our fat compositions can be very low
(ie <5 wt %) we can also make fat compositions with a very good
performance if the trans content is 10-70 wt %. Herefore we can use
partially hardened fats as fat component B or D. However low trans
contents are in general preferred.
Fat B that can be applied in making our fat compositions can be
selected from the group consisting of palm oil mid, palm oil
stearin, cocoa butter and fully or partially hardened vegetable
oils, like fully hardened bean oil, rape seed oil or cotton seed
oil.
Fat C is suitably selected from the group consisting of sunflower
oil, high oleic sunflower oil, olive oil, bean oil, safflower oil,
rape seed oil, palm oil olein, olein fractions of vegetable oils,
high oleic vegetable oils; corn oil; cotton seed oil.
Part of our invention are also the food products with a fat phase
containing or consisting of the fat composition according to our
invention.
Examples of food products, wherein our novel fats can be applied
are: margarines; spreads; ice-creams; confectionery products (in
particular fillings), bakery products (cakes, puff-pastries,
laminated products etc.), cream alternatives, mayonnaises,
dressings, soups and sauces. In these products the following
benefits can be achieved: Higher SV in cakes and laminated
products: higher patty heights in puff-pastries: better aerated
batters for cake: improved overrun and stand-up properties for
cream alternatives, as well as shorter whip-times herefore and
better melt-down and aeration preperties for confectionery
fillings.
EXAMPLES
Methodology
Anti-inflammatory Cell Assays
It is emphasized that the anti-inflammatory effects were determined
by in vitro tests wherein the production of intracellular adhesion
molecules (=ICAM) and Prostaglandin E2 (=PGE2) production by the
human skin fibroblasts is measured after being induced by the
inflammatory modulus phorbyl myristyl acetate (PMA). Effects on
basal PGE2 levels in keratinocytes are also measured. A reduction
of the levels of ICAM and PGE2 is indicative for the
anti-inflammatory effect.
Fibroblast Cell Assay
Primary human foreskin fibroblasts at passage 2 (P2) were seeded
into 96-well plates at 10000 cells/well and maintained for 24 hours
in an atmosphere of 5% carbon dioxide in Dulbeccos Modified Eagles
Medium (DMEM) supplemented with 10% foetal calf serum. Petroselinic
acid was added to fresh cell media (DMEM, supplemented with 10%
foetal calf serum) in dimethylsulphoxide (DMSO, final concentration
1%) in triplicate and incubated for a further 24 hours. Phorbal
myristate acetate (PMA) in ethanol/cell media (10 nm) was added to
the media and the cells incubated for a futher 24 hours. PMA
represents an external stressor which induces oxidative stress and
inflammatory responses in cells. The fibroblasts/media were then
analysed as described below immediately or snap frozen in liquid
nitrogen and stored at -70.degree. C. for future analysis. The
cells were then counted to ensure no effect on cell number.
Prostaglandin E2 (PGE2) Assay
Volumes of 50 .mu.l culture medium were taken for PGE2 assay after
gently shaking the culture plate. PGE2 levels in the medium were
determined with a Biotrak PGE2 immunoassay kit (Amersham, UK). The
assay is based on the competition between unlabelled PGE2 in the
sample and a fixed quantity of horseradish peroxidase labelled PGE2
for a limited amount of fixed PGE2 specific antibody.
Concentrations of unlabelled sample PGE2 are determined according
to a standard curve which was obtained at the same time.
ICAM-1 Assay
Media were discarded and cells washed with Dulbecco PBS. To the
washed cells, 150 .mu.l 0.1% Triton X-100 (Sigma) was added for 3
minutes to extract ICAM from cell membrane. The extracts were
transferred to Eppendoff centrifuge tubes and centrifuged at 1000 g
for 2 min to remove cell debris. A volume of 100 .mu.l supernatant
was used for ICAM assay. The soluble ICAM-1 was assessed with
commercially available immunoenzymometric assay kit (R&D
Systems). Concentrations of ICAM-1 in the samples were determined
based on parallelly running standard curve.
Glutathione (GSH) Depletion Model
The anti-inflammatory effects of petroselinic acid can also be
shown using a glutathione (GSH) depletion model in fibroblasts.
L-Buthionine sulfoximine (BSO, Sigma), a specific inhibitor of
.gamma.-glutamylcysteine synthetase, was used to deplete
intracellular GSH levels. This depletion model is based on the
natural degradation of GSH while synthesis is inhibited. BSO is
added to the fibroblast cell media after 24 hours (0.25 mM), this
is predissolved in DMSO (DMSO, final concentration <1%). In
addition, where stated, the antioxidants Epigallocatechingallate
(ECGC) and Quercetin were also added to the media after 24 h. The
former was dissolved directly in the media, quercetin was initially
dissolved in ethanol, this was then diluted 500 fold into the
media.
Keratinocyte PGE2 Assay
Keratinocytes were grown in 96 well plates to approximately 80%
confluency in keratinocyte growth medium (KGM) which was then
replaced in with KGM without hydrocortisone for 24-48 h. The cells
were then incubated in the presence or absence of petroselinic acid
for 24 h. The medium was then removed and assayed for the
pro-inflammatory PGE.sub.2 content by Enzyme-linked
immunoassay.
Keratinocyte SDS Viability Assay
Keratinocytes were grown in 96 well plates to approximately 80%
confluency in keratinocyte growth medium (KGM) which was then
replaced in with KGM without hydrocortisone for 24-48 h. The cells
were then treated with a concentration of sodium do-decyl sulphate
(SDS) which will produce cell viability of approximately 50% (2
.mu.g/ml). This is done in the presence or absence of petroselinic
acid. After incubating for 24 h the medium was removed and the cell
viability determined by the Neutral Red method. Basically, the
cells were incubated for 3 h in KGM containing 25 .mu.g/ml neutral
red after which the medium was removed and the cells were then
extracted with 1 ml of 1% (v/v) acetic acid, 50% (v/v) ethanol for
30 min. The absorbance of the extract at 562 nm was determined and
the viability evaluated by reference to control wells which
contained neither SDS or petroselinic acid. This methodology has
shown that the keratinocyte toxicity of an irritant relates to the
irritancy effect of the agent in vivo (Lawrence et al Toxicol. In
Vitro 10, 331-340 1996).
Procedure For Measuring Procollagen-I and Decorin Synthesis In
Human Dermal Fibroblasts
Preparation of Dermal Fibroblast Conditioned Medium
Primary human foreskin fibroblasts at passage 2 (P2) were seeded
into 12-well plates at 10000 cells/cm.sup.2 and maintained for 24
hours in an atmosphere of 5% carbon dioxide and 4% oxygen in
Dulbeccos Modified Eagles Medium (DMEM) supplemented with 10%
foetal calf serum. After this time the cells were washed with serum
free DMEM and then incubated in fresh serum free DMEM for a further
60 hours. The fibroblast monolayers were then washed again with
serum free DMEM. Test reagents (petroselinic acid, EGCG and gallic
acid) and vehicle controls were added to the cells in triplicate in
a final volume of 0.4 ml/well fresh serum free DMEM and incubated
for a further 24 hours. This fibroblast conditioned medium was
either analysed immediatedly or snap frozen in liquid nitrogen and
stored at -70.degree. C. for future analysis. The cells were then
counted and data from the dot-blot analysis subsequently
standardised to cell number.
Dot Blot Assay For Decorin Protein in Dermal Fibroblast Conditioned
Medium
Samples of conditioned medium from dermal fibroblasts treated with
vehicle (as a control) or test reagents were supplemented with 20
mM dithiothreitol (1:10 dilution of 200 mM stock solution) and 0.1%
sodium dodecylsulphate (1:100 dilution of 10% stock solution),
mixed well and then incubated at 75.degree. C. for 2 minutes. A
standard for the assay was generated by serial dilution of neat
fibroblast conditioned medium from fibroblasts seeded at 10000
cells/cm.sup.2 in a 175 cm.sup.2 flask and maintained in serum free
DMEM as described above. Assay samples were subsequently applied in
triplicate to a prewetted sheet of Immobilon-P transfer membrane
using the 96-well Bio-Dot Apparatus from Bio-Rad as described in
the manufacturers guidelines. Approximately 200 .mu.l of medium was
applied per well. The medium was allowed to filter through the
membrane under gravity (30 minutes) after which the membrane was
washed twice with PBS (200 .mu.l). These PBS washes were allowed to
filter through the membrane under gravity (2.times.15 minutes). The
Bio-Dot apparatus was then attached to a vacuum manifold and a
third and final PBS wash carried out under suction. The apparatus
was disassembled, the membrane removed and quickly cut as required
before being placed in blocking buffer overnight at 4.degree. C.
Membranes prepared for decorin analysis were blocked with 3% (w/v)
bovine serum albumin (BSA)/ 0.1% (v/v) Tween 20 in phosphate
buffered saline (PBS), whilst those for procollagen-I analysis were
blocked with 5% (w/v) non fat dried milk powder/0.05% Tween 20 in
PBS. The following day, the membranes were probed with 1:10000
dilution of primary antibodies to human decorin (rabbit polyclonal;
Biogenesis) for 2 hours at room temperature. The membranes were
subsequently washed with TBS/0.05% Tween 20 (3.times.5 minutes) and
then incubated with 1:1000 dilution of .sup.125 I-conjugated
anti-rat or anti-rabbit F(ab')2 fragments (Amersham) as required
for 1 hour at room temperature. Following this the Immobilon strips
were again washed with TBS/Tween 20 (3.times.5 minutes) before
being allowed to dry in air at room temperature. The dried
membranes were wrapped in cellophane and exposed to a Molecular
Dynamics storage phosphor screen for 16-18 hours. At the end of
this time the exposed screen was scanned by a phosphorimager
(Molecular Dynamics Phosphorimager SF) using ImageQuant.TM.
software. Dot intensity was assessed by computer-assisted image
analysis using the quantification tools in ImageQuant.TM.,
standardised to cell number and the effects of various test
reagents on decorin and procollagen-I synthesis were determined
relative to a vehicle treated control value of 100 arbitrary
units.
EXAMPLES
1. Anti-inflammatory Effects in Fibroblasts
FIG. 1 demonstrates that challenging cells with an inflammatory
stimulus such as PMA (Phorbol myristyl acetate) causes an increase
in the inflammatory response as measured by prostaglandin E2 (PGE2)
production. Petroselinic acid, even at the levels of 0.1 .mu.M,
dramatically reduces the inflammatory response as measured by PGE2
production. --good anti-inflammatory activity.
FIG. 2 demonstrates that petroselinic acid also decreases the
production of Intracellular adhesion molecule (ICAM) in
fibroblasts, which is another marker of inflammation. The reduction
in ICAM occurs under both basal condition (control) and in cells
stimulated with an inflammatory stimulus (in this case PMA).
The data for FIG. 1 represents the mean.+-.standard deviation
(n=3). The experiment being performed in triplicate to confirm the
effect. The levels of PGE2 were measured in untreated cells
(Control), cells treated with the inflammatory stimulus PMA (PMA)
and PMA treated cells pre-incubated with 0.1, 1.0, 10, and 100
.mu.M petroselinic acid. A significant decrease in PGE2 response
was found for petroselinic acid treated cells.
The data for FIG. 2 represents the mean.+-.standard deviation
(n=3). The experiment being performed in triplicate to confirm the
effect. The levels of ICAM were measured in untreated cells
(Control), petroselinic acid treated cells (Petroselinic acid 100
.mu.M), cells treated with the inflammatory stimulus PMA (PMA) and
PMA treated cells pre-incubated with 0.1, 1.0, 10, and 100 .mu.m
petroselinic acid. A significant decrease in ICAM response was
found for petroselinic acid treated cells.
2. Anti-inflammatory Effects--Synergy With Antioxidants
FIG. 3 demonstrates the effects of BSO and PMA on inflammation as
measured by the levels of ICAM. Antioxidants decrease the ICAM
response at a concentration of 10 .mu.M quercetin and ECGC. In this
case petroselinic was added to the media at the lower level of 0.01
.mu.M and had no effect. Yet in combination, petroselinic acid had
a synergistic effect with the antioxidants tested.
The data for FIG. 3 represents the mean.+-.standard deviation
(n=3). The experiment being performed in triplicate to confirm the
effect. The levels of ICAM were measured in untreated cells
(Control), BSO plus PMA treated cells (BSO&PMA), and cells
treated with the inflammatory stimulus PMA and BSO pre-incubated
with; A: petroselinic acid (0.1 .mu.M), B: EGCG (10 .mu.M), C:
quercetin (10 .mu.M), D: petroselinic acid plus EGCG (0.1 .mu.M),
E: petroselinic acid plus quercetin (0.1 .mu.M+10 .mu.M). A
significant and synergistic decrease in ICAM response was found for
petroselinic acid treated cells in the presence of
antioxidants.
The data for FIG. 4 represents the mean.+-.standard deviation
(n=3). The levels of ICAM were measured in untreated cells
(Control), PMA treated cells (PMA), and cells treated with the
inflammatory stimuli PMA pre-incubated; petroselinic acid (1
.mu.M), genistein (1 .mu.M) and petroselinic acid plus genistein (1
.mu.M) each). A significant and synergistic decrease in ICAM
response was found.
3. Anti-inflammatory/Anti-irritancy Effects in Keratinocytes
Petroselinic acid was shown to significantly reduce the basal
levels of secreted PGE.sub.2 in keratinocytes, indicative of
reduced inflammatory potential (FIG. 5). In addition, treatment
with petroselinic acid reduces the toxic effects of SDS on
keratinocytes indicating that it will reduce skin irritancy (FIG.
6).
The data for FIG. 5 represents the mean.+-.standard deviation
(n=4). The levels of PGE2 were measured in untreated cells (Co),
and petroselinic acid treated cells (Pet 0.1 .mu.M and 1 .mu.M).
The application of petroselinic acid reduced the basal levels of
secreted PGE.sub.2 indicative of reduced inflammatory potential.
Statistical comparison of control (Co) and 1 .mu.M petroselinic
acid showed p<0.05 by Student's t-test.
The keratinocyte viability (FIG. 6) was measured in response to SDS
(2 .mu.g/ml) in the presence of petroselinic acid (0.01, 0.1, 1, 10
.mu.M) and data represented as a percentage of the control (no
petroselinic acid). All petroselinic acid values were significantly
increased compared to the control as determined by Iway ANOVA with
Student-Neumann-Kuels multiple comparison, p<0.05.
4. Anti-ageing Effects--Determined By Measurement of Decorin Levels
and Synergy With Antioxidants
Petroselinic acid has been demonstrated to boost decorin levels in
fibroblasts, consistent with dermal repair and antiageing activity
(Table 1). Antioxidants such as ECGC, gallic acid, diadzein and
genistein can also exert this effect (Table 2). In combination, a
synergistic boost in decorin levels is observed (Table 3).
Petroselinic acid (.mu.M) Decorin (% of control) 0 100.0 .+-. 13.6
1 137.6 .+-. 13.3 10 152.5 .+-. 14.2
TABLE 1 Boosting of decorin by petroselinic acid. The production of
decorin by fibroblasts was determined in control cells (no
treatment) and petroselinic acid treated cells. The data is
represented mean +/- standard deviation as decorin levels as a
percentage of the control (no petroselinic acid). Each data point
was determined in triplicate. All petroselinic acid values were
significantly increased compared to the control. Antioxidants
Decorin (% of control) EGCG (.mu.g / ml) 0 100.0 .+-. 6.4 1 167.5
.+-. 11.8 2.5 188.6 .+-. 9.4 5 258.2 .+-. 6.1 Gallic acid (.mu.g /
ml) 0 100.0 .+-. 7.0 1 136.9 .+-. 4.2 2.5 163.4 .+-. 6.5 5 161.8
.+-. 6.6
TABLE 1 Boosting of decorin by petroselinic acid. The production of
decorin by fibroblasts was determined in control cells (no
treatment) and petroselinic acid treated cells. The data is
represented mean +/- standard deviation as decorin levels as a
percentage of the control (no petroselinic acid). Each data point
was determined in triplicate. All petroselinic acid values were
significantly increased compared to the control. Antioxidants
Decorin (% of control) EGCG (.mu.g / ml) 0 100.0 .+-. 6.4 1 167.5
.+-. 11.8 2.5 188.6 .+-. 9.4 5 258.2 .+-. 6.1 Gallic acid (.mu.g /
ml) 0 100.0 .+-. 7.0 1 136.9 .+-. 4.2 2.5 163.4 .+-. 6.5 5 161.8
.+-. 6.6
TABLE 3 Synergistic boosting of decorin by petroselinic acid and
antioxidants The production of decorin by fibroblasts was
determined in control cells (no treatment) and petroselinic acid
and/or antioxidant treated cells. The data is represented as mean
+/- standard deviation (n = 3) for decorin levels as a percentage
of the control (no antioxidant). A synergistic effect in boosting
decorin levels of both antioxidants and petroselinic acid was
observed, compared to the control.
5. 80% Fat Examples
Three margarines were produced using exactly the same process
conditions.
A. Formulation Aqueous Phase Water 18.48% Potassium Sorbate 0.15
Citric Acid 0.07 SMP 1.0 Fat Phase Fat Blend 80.0 Hymono 8903 0.3
Fat Phase: Product 1. 13% InEs, 87% SF (Control) Product 2. 13%
InEs, 12% Coriander Oil, 75% SF (=according to EP777971) Product 3.
13% InEs, 47% Coriander Oil, 40% SF InEs = interesterified palmoil
58/palm kernel olein ratio 1:1 Coriander: contained 46 wt % PSA. SF
= sunflower oil
B. Process Conditions
The process line was configured as:
Premix temperature was set at 65.degree. C. and 60-rpm stirrer
speed. All units were set to 15 AC, with shaft speeds set to 1000
rpm. Throughput was 50 g/min. using the constant displacement pump.
A coarse premix was prepared by slowly adding the prepared aqueous
phase to the oil phase in the premix tank. A 2 kg-batch size was
employed. The mix was allowed to stir for 15 minutes before pumping
was commenced. After pumping was started, the line was allowed to
run for 15 minutes before any collection of product.
The following process parameters were noted:
A.sub.1 exit C.sub.1 exit A.sub.2 exit Line Pressure Product
(.degree. C.) (.degree. C.) (.degree. C.) (bar) 80% Fat Control
20.1 18.4 18.4 1.5 80% Fat, 12% 20.6 18.5 18.4 1.3 Coriander 80%
Fat, 47% 20.8 18.6 18.4 1.4 Coriander
Five tubs of each product were collected and placed at 5.degree. C.
After one day, one tub of each was transferred to each of
5.degree., 10.degree., 15.degree. and 20.degree. C. for evaluations
after one week.
6. Product Assessment
Product 3 (47% Coriander) was pale yellow in colour, with the
others being very white.
All samples spread easily and smoothly, with no water loss evident.
No difference in taste was found between samples. Good breakdown in
the mouth was evident for all samples.
C-Value Collar Conductivity Sample (g/cm.sup.2) (Scale I to VI)
(.mu.Scm.sup.-1) 5.degree. C. Storage 1. 80% Fat Control 725 II
<10.sup.-5 2. 80% Fat, 12% 675 II <10.sup.-5 Coriander 3. 80%
Fat, 47% 750 II <10.sup.-5 Coriander 10.degree. Storage 1. 80%
Fat Control 580 II <10.sup.-5 2. 80% Fat, 12% 560 II
<10.sup.-5 Coriander 3. 80% Fat, 47% 590 II <10.sup.-5
Coriander 15.degree. C. Storage 1. 80% Fat Control 380 I
<10.sup.-5 2. 80% Fat, 12% 360 I <10.sup.<5 Coriander 3.
80% Fat, 47% 410 I <10.sup.-5 Coriander 20.degree. C. Storage 1.
80% Fat Control 360 I <10.sup.-5 2. 80% Fat, 12% 340 I
<10.sup.-5 Coriander 3. 80% Fat, 47% 330 I <10.sup.-5
Coriander
C-value Measurement
C-values are determined using a cone penetrometer (manufactured by
SUR) and a 40.degree. cone (weight 80 g). The point of the cone is
placed just in contact with the surface of the spread and the cone
allowed to drop for 5 seconds. The distance travelled is thus
proportional to the hardness of the product.
The C-value can then be calculated from C=K.F/P.sup.1.6
Where
C=Yiel or C-value in (g/cm.sup.2)
F=Total weight of cone and sliding stem (g)
P=Penetration depth (0.1 mm)
K=Fatctor for cone angle (40.degree.=5840)
Conductivity
Conductivity is measured using a cell manufactured by URL
Vlaardingen consisting of two parallel plates 1 cm apart,
approximately 3.times.2 cm in size. This is connected to a Philips
PW 9526 digital conductivity meter. Conductivity can then be
directly read off (in .mu.Ssm.sup.-1) after inserting the probe
into the spread.
Collar
Collar is measured by inserting a 5 mm diameter steel rod to a
depth of 10 mm into the spread. The resulting raising of spread
around the rod is graded on a scale of I to VI, with I meaning no
raising of product around the rod. VI indicating large amounts of
spread raised and cracking of the product around the rod.
7. 40% Fat Examples
Two halvarines were produced, one with coriander oil and a control
without.
a. Formulation Aqueous Phase Water 56.48% Potassium Sorbate 0.15
Citric Acid 0.07 Gelatine (240 1.5 bloom) SMP 1.5 Fat Phase Fat
Blend 40.0 Hymono 7804 0.3 Fat Phase: Product 1. 13% InEs, 87% SF
(Control) Product 2. 13% InEs, 42% Coriander Oil, 45% SF
b. Process Conditions
The process line was configured as:
Premix temperature was set at 65.degree. C. and 60-rpm stirrer
speed. A-units were set to 10.degree. C., with shaft speeds set to
1000 rpm. C-units were set at 15.degree. C., also with shaft speeds
of 1000 rpm. Throughput was 30 g/min. using the constant
displacement pump. A coarse premix was prepared by slowly adding
the prepared aqueous phase to the oil phase in the premix tank. A 2
kg-batch size was employed. The mix was then allowed to stir for 15
minutes before pumping was commenced. After pumping was started,
the line was allowed to run for 15 minutes before any collection of
product.
The following process parameters were noted:
Line A.sub.1 exit C.sub.1 exit A.sub.2 exit C.sub.2 exit Pressure
Product (.degree. C.) (.degree. C.) (.degree. C.) (.degree. C.)
(bar) 40% Fat Control 21.1 16.5 18.3 13.8 3.5 40% Fat, 42% 22.6
17.3 18.5 13.9 3.8 Coriander
Five tubs of each product were collected and placed at 5.degree. C.
After one day, one tub of each was transferred to each of
5.degree., 10.degree., 15.degree. and 20.degree. C. for evaluations
after one week.
C. Product Assessment
The 42% Coriander sample was slightly more yellow in colour than
the control, but not as marked a difference as with the
margarine.
All samples spread easily and smoothly, with no water loss evident.
No difference in taste was found between samples. Good breakdown in
the mouth was evident for all samples. Improved hardness was found
at 5.degree. C., this is beneficial to make low fat products with
reduced levels of hardstock.
C-Value Collar Conductivity Sample (g/cm.sup.2) Scale I to VI)
(.mu.Scm.sup.-1) 5.degree. C. Storage 1. 40% Fat Control 410 II
<10.sup.-4 2. 40% Fat, 42% 600 II <10.sup.-4 Coriander (on
fat) 10.degree. Storage 1. 40% Fat Control 400 II <10.sup.-4 2.
40% Fat, 42% 410 II <10.sup.-4 Coriander (on fat) 15.degree. C.
Storage 1. 40% Fat Control 240 I <10.sup.-4 2. 40% Fat, 42% 250
I <10.sup.-4 Coriander (on fat) 20.degree. C. Storage 1. 40% Fat
Control 230 I <10.sup.-4 2. 40% Fat, 42% 250 I <10.sup.-4
Coriander (on fat)
8. Ice Cream
The ice creams were made according to the following recipes.
Recipe: Wt % Fat blend 10 Skimmed 10 milkpowder Icing sugar 12 Corn
syrup 4 solids Dextrose 2 monohydrate Dimodan PVP 0.6 Water
61.4
The fat blend for the reference nol was 30% POf IV65/20% CN/50% SF
and the fatblend according to the invention no 2 was 30% POf
IV65/20% CN/50% Coriander.
POf IV 65=palm olein Iodine value: 65
CN=coconut oil
Processing
All ingredients except the water and the fat were mixed. Then the
cold water was added to this mixture. This mixture was heated in a
water bath to a temperature of 70.degree. C. Then the fully liquid
fat blend comprising the palm oleine (PO-f), coconut oil (CN) and
sunflower oil (SF) (=reference) or Coriander oil (blend 2) was
added to the mixture and was stirred in the ultra-turrax. This
emulsion was cooled in a water bath of 20.degree. C. The emulsion
was stirred in the ultra-turrax again. The batch ice cream machine
was held for 24 hours at -28.degree. C. prior to use. The emulsion
was placed in the batch ice cream machine and was stirred for 15
minutes. The resulting ice cream was stored at -18.degree. C. for 3
days and was then evaluated.
Evaluation
The following evaluations were performed:
Processing of the materials
Viscosity measurement of the ice cream emulsion prior to freezing
at 20.degree. C. Method: Haake viscometer using Casson regression
method.
Overrun directly after freezing. Method: Standardized amount of
product is weighed before and after freezing/aeration process to
determine amount of air incorporated into the product during
freezing/aeration process (overrun measured in % air on total
weight)
Hardness of the ice cream after storage for 3 days in the freezer
Method: Stevens texture analyzer using 60.degree. cone, Speed 0.5
mm/sec and penetration depth 2 mm. Hardness measured in grams
Sensory evaluation (hardness, texture, melt down and flavor
release)
Results ice cream preparation
Both ice creams could be processed using standard equipment.
1 2 Viscosity ice at 20.degree. C. Eta (Pas) 0.008174 0.009123 Tau0
(Pa) 0.111 0.05834 Cupweight at 20.degree. C. 70.1 73 (g) Cupweight
Cupweight Temp (.degree. C.) (g) Temp (.degree. C.) (g) 0 min 6.1
94.8 6.3 92.9 20 min -4.5 87.4 -4.5 88.4 3 days freezer -18 86.5
-18 86.7 Overrun % 9.6 7.2
Hardness after 3 days in the freezer
1 2 76 90
Sensory Evaluation
The two ice creams were evaluated by the taste panel after 2 weeks
storage at -18.degree. C. The results were as follows:
Hardness: the sample was slightly harder than the reference
Meltdown was similar
Texture: Sample was slightly more grainy, and stayed longer
firm.
In general the ice cream samples were of good quality. They were
firm, had good texture and oral melt.
9. Bakery Application
Experimental
Reference for bakery application is "Biskien Zacht". The following
blends were made:
Blend: 1 2 3 Ref SHs 30 40 50 POf IV65 25 20 15 Coriander oil 45 40
35 Biskien zacht.sup.R 100 Ref NMR us N10 27.8 37.2 47.5 58 N20
17.7 26.9 35.6 36 N25 8.3 17 25.5 25 N30 2.8 7.7 14.4 14 N35 0.5 1
1.4 5
SHs=shea stearin
The reference and the best match (blend 3)were evaluated in a
standard biscuit recipe.
Recipe
1 Biskien zacht.sup.R
2 SHs/POfIV65/Coriander oil 50/15/35
(%) Water 5.12 Fat 29.0 Sugar 23.9 Sodiumbicarbonate 0.34 Lemon
peel 0.68 Flour 41 100
Processing
The dough was prepared in advance and kept in the refrigerator for
1 night according to the following procedure.
Sugar and fat were mixed to smooth mass and then sugar, lemon peel
and flour were added to this mixture. Finally the water was
added.
The biscuits were baked during 18 minutes at 155.degree. C.
Evaluation
Dough could be processed using standard equipment and the product
had acceptable sensory properties.
10. Confectionery
Recipe
The following recipe was used for the evaluation of the fat blends
in a filling application:
Fat blend 35 Cocoa powder 10 Skimmed milk powder 7 Sugar 48
Lecithin 0.5
The blends to be evaluated were:
1. 40/10/50 CCBs/POfIV65/SF (Reference)
2. 40/10/50 CCBs/POfIV65/Coriander oil
3. 40/60 CCBs/Coriander oil
CBs is cocoa butter stearine
Processing
The fillings were prepared using roller refiner and conche. The
fillings were cooled to 29.degree. C. before depositing in
aluminium cups.
The Fillings Were Evaluated On
Hardness was measured by use of the Stevens Texture Analyser after
(STA) 3 days at 20.degree. C., Cone 60.degree., penetration 2
mm:
Fat type STA (g) 1 10/10/50 CCBs/POfIV65/SF (Ref) 179 2 40/10/50
CCBs/POfIV65/Coriander 288 oil 3 40/60 CCBs/Coriander oil 344
Sensory Properties
Sample 2 looked like the reference and was slightly softer. Sample
1 and 2 were more chewy and more plastic. Sample 3 was harder, less
plastic, not chewy and melted quicker and cooler. This sample was
less waxy and had slightly more viscosity of melt.
Composition of Fat Blends Used
Components SAFA Unsat. Sym Unsat. asym POf 40 60 0 CN 94 6 0 SF 13
87 0 Coriander 5 33 60 SHs 64 36 0 CCBs 66 34 0 Biskien n.m. n.m. 0
zacht
Ex 8.1=30/20/50 POf/CN/SF
Ex 8.2=30/20/50 POf/CN/Coriander oil
Ex 9.1=Biskien zacht.sup.R
Ex 9.2=50/15/35 SHs/POf/Coriander oil
Ex 10.1=40/10/50 CCBs/POf/SF
Ex 10.2=40/10/50 CCBs/POf/Coriander oil
Ex 10.3=40/60 CCBs/Coriander oil
Unsat. SAFA Sym Unsat asym 8.1 37.3 62.7 0 8.2 33.3 35.7 30 9.1
n.m. n.m. 0 9.2 39.75 38.55 21 10.1 36.9 63.1 0 10.2 32.9 36.1 30
10.3 29.4 33.4 36
* * * * *